We have been studying the effect of DNA and RNA secondary structure on gene expression, DNA replication and mutation. In particular we have been examining the role of unusual nucleic acid structures in the etiology of fragile X syndrome as well as a number of other genetic defects. Fragile X syndrome is the most common cause of inherited mental retardation, occurring at a frequency of about 1 in 1500 males and 1 in 2500 females. Most mutations associated with this syndrome coincide with gaps of poorly staining chromatin on the X chromosome seen when cells containing this mutation are starved for the precursors of DNA synthesis. These so-called fragile sites are associated with a sudden, large increase in the number of repeats of the triplet CGG found in the 5' untranslated region of a gene called FMR-1. This so-called triplet expansion results in increased methylation of the FMR-1 promoter and the absence of detectable mRNA. Six other triplet expansion diseases are currently known, that are due either to expansion of CGG-repeats, or to the expansion of a repeat with the sequence CAG. We have shown that CGG-repeats can form a number of unusual nucleic acid structures, including an unusual hairpin in the absence of K+, and an intrastrand tetraplex containing both guanine tetrads and C.C basepairs in the presence of this cat ion. The tetraplex forms a strong block to DNA synthesis that might provide a biological rationale for triplet expansion by promoting repeated strand slippage during replication. We have shown that similar structures are also found associated with other hypervariable sequences where it is possible that they affect DNA stability in much the same way. Other work in progress in this group includes a study of a conserved RNA structure found in non-LTR retrotransposons in mammals, a study of an unusual DNA structure we call a cinched tetraplex in the promoter of the chicken beta-globin gene, and a study of the effect of triplex formation on transcription in vitro and in vivo.